Allele-specific amplification of nucleic acids allows for simultaneous amplification and analysis of the target sequence. Allele-specific amplification is commonly used when the target nucleic acid has one or more variations (polymorphisms) in its sequence. Nucleic acid polymorphisms are used in DNA profile analysis (forensics, paternity testing, tissue typing for organ transplants), genetic mapping, distinguishing between pathogenic strains of microorganisms as well as detection of rare mutations, such as those occurring in cancer cells, existing in the background of cells with normal DNA.
In a successful allele-specific amplification, the desired variant of the target nucleic acid is amplified, while the other variants are not, at least not to a detectable level. A typical allele-specific amplification assay involves a polymerase chain reaction (PCR) with at least one allele-specific primer designed such that primer extension occurs only when the primer forms a hybrid with the desired variant of the target sequence. When the primer hybridizes to an undesired variant of the target sequence, primer extension is inhibited.
Many ways of enhancing allele-specificity of primers have been proposed. However, for many clinically-relevant nucleic acid targets lack of specificity of PCR remains a problem. Therefore radically novel approaches to design of allele-specific primers are necessary.